Expression levels, transcript abundance (TA), of genes in peripheral blood cells are important biomarkers. Current methods using gene TA in peripheral blood samples as biomarkers are performed in a cell mixture samples of peripheral blood in such a way that no information can be obtained about the TA of a specific hematopoietic subpopulation, such as B cell lymphocytes. For example, some methods obtain expression level results of all cell types in the cell mixture, and not that of a specific hematopoietic subpopulation.
Specifically, in the mathematical method of deconvolution (Lu et al.), the fractional proportion of each cell types is determined in a cell-mixture sample. The expression level of a gene in a particular cell type in a group of samples is assumed to be a constant. The expression in a cell-mixture sample is simply determined based on the fractional proportion of a specific subpopulation in the sample, and is not a measure of the actual expression in the subpopulation of an individual, as it is assumed to be constant.
On the other hand, gene TA of specific hematopoietic subpopulation (e.g., a leukocyte subpopulation) are the preferred biomarkers. In order to obtain TA of a specific hematopoietic subpopulation, the current state of the art requires a prior cell separation or cell sorting to isolate the specific hematopoietic subpopulation from peripheral whole blood sample before quantitative analysis of gene expression. Cell subpopulations separation is laborious and tedious. Such procedure is basically not practical in the setting of a clinical service laboratory. Thus, there are limitations in current methods of analysis of gene expression in peripheral blood samples.
Extensive research has been carried out to study the gene expression level in purified hematopoietic (specifically leukocyte) subpopulations. They are carried out after procedures of cell separation or cell culture in order to obtain samples of a specific leukocyte subpopulation that is under research. There has also been a strong interest to examine the gene expression levels in different types of peripheral blood samples, including whole blood and peripheral blood mononuclear cells samples as biomarkers of diseases, treatment response, and prognostic markers.
Therefore, it is desirable to provide new methods, apparatuses, systems, and compositions to eliminate the requirement of cell purification or separation in the process to determine the levels of gene expression or TA of target genes in a specific hematopoietic subpopulation (e.g., leukocyte cell-type), as well as provide other advantages.